This mentored K23 proposal outlines the 4 year training program for Dr. Kashyap in the area of endothelial function in peripheral arterial disease (PAD). The applicant has a strong background in vascular disease and thrombus-endothelial interaction. The long-term objective of the applicant is to understand the effect of endothelial function on atherogenesis, ischemic clinical events, and durability of endovascular intervention in PAD. This will be accomplished with a program of didactics, mentored training in clinical research, and training in clinical trial methodology. The candidate has made the persistent observation that thrombus causes endothelial dysfunction. Previous experiments and preliminary data indicate that luminal arterial thrombus decreases nitric oxide (NO) by increasing arginase activity. Removal of the thrombus by mechanical means or dissolution with thrombolysis restores blood flow but leads to persistent endothelial dysfunction. In animal models, thrombolysis coupled with NO's precursor, L-arginine, ameliorates thrombus induced endothelial dysfunction. The hypotheses to be tested are A) Acute arterial thrombosis worsens endothelial dysfunction by increasing arginase activity and B) Endothelial function after thrombosis can be improved with regional L-arginine supplementation by increasing local nitric oxide levels. The interrelated specific aims for this proposal are: 1. Assess endothelial function in human arteries from fresh amputation specimens and correlate with arginase activity, NO, and arterial morphology. 2. Assess endothelial function with Intravascular Ultrasound (IVUS) in vivo at the time of angiography for lower extremity PAD. 3. Compare L-arginine to placebo in a prospective, double-blind study in patients with critical limb ischemia requiring thrombolysis. The results of these studies will lead to a better understanding of endothelial function directly at the sites of peripheral arteries affected by atherosclerosis. This insight will lead to targeted therapies to halt or improve endothelial dysfunction. The proposed studies will impact patients with PAD and improve clinical care of patients with critical ischemia. Lay Summary: Narrowing in the leg arteries of humans can lead to clotting of the arteries and amputation. When clots form, the cells lining of the arteries do not work correctly and can cause re-clotting despite using clot-dissolving agents. We will test a novel clot-dissolving combination that may improve cellular function and outcomes after artery clotting.